Indian Journal of Animal Research

  • Chief EditorK.M.L. Pathak

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Haematological and Serum Biochemical Changes in Chronic Laminitis Affected Horses

K.C. Ogbanya1,*, C.A. Eze1, J.I. Ihedioha2
1Department of Veterinary Surgery, University of Nigeria, Nsukka, Nigeria.
2Department of Veterinary Pathology and Microbiology, University of Nigeria, Nsukka, Nigeria.
Background: Equine laminitis is considered as one of the major health problems affecting equine species. Haematological and serum biochemical assessments are often used for diagnosis of both human and animal diseases. Despite numerus findings on induced acute laminitis in horses, little is known about the haematological and serum biochemical changes associated with chronic laminitis. The current study aimed to evaluate the hematological and biochemical changes in horses with chronic laminitis.

Methods: A total of 60 horses comprising of 22 apparently healthy horses (group A) and 38 horses radiographically diagnosed as laminitis (group B) were studied during 2018-2019 laboratory investigation. Blood collection for hematology and serum biochemistry were carried out aseptically via the jugular vein. The packed cell volume (PCV) and hemoglobin concentration (Hb) were determined using micro-haematocrit and cyanomethaemoglobin methods respectively. White blood cell (WBC) and red blood cell (RBC) counts were determined using haemocytometer method. Serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities were determined by the Reitman-Frankel method. Serum alkaline phosphatase (ALP) activity, total protein, albumin and creatinine were determined using phenolphthalein monophosphate, biuret, bromocresol green and modified Jaffe methods respectively. Parameters were analyzed using student t-test. 

Results: The PCV, HB, RBC, WBC and neutrophil counts in groups B were significantly lower compared to group A (control). Lymphocytes, monocytes, basiophils, eosinophils and the biochemical parameters in the control group (group A) horses did not vary significantly (P > 0.05) with group B (horses with laminitis). The haematological profiles depict anaemic state of the laminitis in horses. Non-differing enzymatic activities in the two groups could suggest that there may not be muscular, renal, intestinal and hepatic disorders involvement with the occurrence of chronic laminitis in the assessed horses.
Equine laminitis is a degeneration of the sensitive laminae of the hoof caused by different factors such as, trauma, increased weight bearing on one limb, excessive work on hard surface, infection, intoxication and overeating of grains and feeding on lush pasture rich in fructan (Radostitis et al., 2007). The basic lesion of laminitis is the separation of the sensitive laminae of the third phalanx from the interdigitating laminae lining the internal surface of the hoof, so that the third phalanx drops through the hoof and comes to rest on the sole (French and Pollitt, 2004). Laminitis is a major cause of lameness in equine species and clinically manifest as alteration in the normal gait (Singh et al., 2015). Lameness is considered to be the third most expensive health problem in animals, having adverse effect on their welfare (Singh et al., 2015).

Laminitis is a metabolic disease of great complexity resulting in diminution of blood perfusion in the capillary of the foot (Riber et al., 1995). It has been demonstrated as being a highly complex metabolic disease associated with disorders of the cardiovascular system, the endocrine system and causing acid-basic and coagulation imbalance (Riber et al., 1995).

Biochemical and hematological parameters may act as a key for the diagnosis of various diseases in both human and animals (Kumar et al., 2019). Evaluation of the blood is commonly used for diagnosis of various diseases of animals, including horses (Schalm et al., 1975; Radostits et al., 1994). A good way of detecting health problems is through a comprehensive evaluation of the blood profile: a complete blood count and serum biochemical evaluation (Hussein and Al-Derawie, 2012). An evaluation of the serum biochemistry is important because of the predictive value of serum biochemical alterations in the assessment of pathological changes in vital internal organs of the body such as liver, kidney, pancreas, heart and muscles (Tyson and Sawhney, 1985; Coles, 1986; Stockham and Scott, 2008). An assessment of the haematological and biochemical changes of horses suffering from chronic laminitis is proper for the understanding of the pathogenesis which will be very useful in formulating the proper therapeutic measures (Kumar et al., 2006).

Induced acute laminitis in horses has been studied and reported to be associated with changes in some haemotological and serum biochemical changes (Kameya, 1973; Riber et al., 1995; Hussein and Al-Derawie, 2012 and Sharma et al. 2015).
A total of 60 horses comprising of 22 apparently healthy horses (group A) and 38 radiographically confirmed chronic laminitis horses (group B) were selected from Obollo-Afor Horse Depot and studied. The study was carried at the Veterinary Teaching Hospital, University of Nigeria, Nsukka between September 2018 to December 2018. Haematological and serum biochemical characteristics of the horses were evaluated and compared between groups (A and B). Five millilitres (5ml) of blood was collected aseptically from the jugular vein of each of the 60 horses in the mornings with the assistance of attendants. Two millilitres from the 5ml was quickly dispensed into bottles with ethylene diaminetetracetate (EDTA) for the haematological evaluation. The remaining 3ml of blood was dispensed into a plain glass test tube and allowed to clot at room temperature. The serum for biochemical determinations was separated from the clot within one hour of blood collection by centrifugation at 3,000 revolutions per minute for 10 minutes using a table centrifuge (TDL4®, B. Bran Scientific and Instruments Co., England).

The packed cell volume (PCV) was determined by the micro-haematocrit method (Thrall and Weiser, 2002). The haemoglobin concentration (Hb) was determined by the cyanomethaemoglobin method (Higgins et al., 2008). The red blood cell (RBC) and total leukocyte counts (TLC) were done by the haemocytometer method, while thin blood smear made on clean grease-free glass slides for differential leukocyte count were stained following the Leishman technique and enumerated by the meander counting method (Thrall and Weiser, 2002).

The serum biochemistry determinations followed standard procedures. QuimicaClinicaAplicada (QCA) test kits (QCA, Spain) were used for all the serum biochemistry determinations.The serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities were determined by the Reitman-Frankel method (Reitman and Frankel, 1957), while the serum alkaline phosphatase (ALP) activity was determined by the phenolphthalein monophosphate method (Klein et al., 1960; Babson et al., 1966). The serum total protein was determined by the direct Biuret method (Lubran, 1978), while the serum albumin was determined by the bromocresol green method (Doumas et al., 1971) and serum globulin was calculated as the difference between the serum total proteins and serum albumin (Johnson, 2008). The serum creatinine was determined by the modified Jaffe method (Blass et al., 1974).

The SPSS statistical package was used to analyze data generated from the study. The haematological and serum indices in the two groups were subjected to student t-test and variant means were separated by the least significant difference method. Significance was accepted at p < 0.05.
The results of the hematological indices were summarized in Table 1 while the serum biochemistry profiles were summarized in Table 2.  Gross photographic illustrations of hoof with chronic laminitis is shown in Fig 1. Radiographic illustration of hooves with chronic laminitis are showed in Fig 2 and 3.

Table 1: Comparative mean value of haematological parameters of apparently healthy (group A) and horses with laminitis (group B).

Table 2: Comparative mean values of serum biochemistry profile of apparently healthy horses (group A) and horses with laminitis (group B)

Fig 1: Photographic picture of a hoof with laminitis showing serous fluid exudation (arrows) which are characteristics of laminae and dorsal coronary band compromise.

Fig 2: Lateromedial radiograph of hoof with laminitis as evidenced in the increased proximal-hoof phalanx distance (A) and distal-hoof phalanx distance (B).

Fig 3: Lateral radiograph of a hoof with laminitis as evidenced by the presence of with Founder Distance (FD) (increased coronary extensor distance (CED))

The mean values of PCV, HB and RBC were significantly lower (p < 0.05) compared to the apparently healthy horses (group A). Similar significant lower WBC counts in group B horses compared to the apparently healthy horses (group A) was recorded. The observed significant decrease in PCV, HB and RBC in chronic laminitis affected horses is similar to the findings of Sharma et al. (2015) who reported a decrease in the PCV and RBC in chronic cattle laminitis. This significantly lowered PCV, HB and RBC depicts the anaemic state of the laminitis in horses. This decreased effects may be associated with the pathogenesis of laminitis which is often characterized with acute or chronic inflammation of the laminae (Sharma et al., 2015). The most common cause of low RBC level in the circulation is chronic inflammation. Chronic inflammation is commonly related to conditions such as abdominal or hoof abscesses or laminar damage, cellulitis, pneumonia and peritonitis (Stockham and Scott, 2008). This happens because the body sequesters iron, which is necessary for red blood cell production, away from the infected area. The significant decrease in the PCV, HB and RBC in the horses with laminitis could also be due to infection condition, or immune mediated diseases. Nutritional factor (nutritional anaemia) could also have predisposed the horses to laminitis since lame horses may not be able to walk around and feed adequately. These findings are however not in agreement with those of Kameya (1973) and Riber et al., (1995), who reported non-significant differences in RBC values of healthy and acute laminitis affected horses. The differences in this study with report of Kameya (1973) and Riber et al., (1995) could stem from the fact that this study was carried out in chronic laminitis whereas their results were from acute induced laminitis. The discrepancies between this result and the previous findings could also be due to variation of etiology of the laminitis.

With regards to differential leucocytes (WBC) counts, a significant (p < 0.05) decrease in the mean values of neutrophil was found in the chronic laminitis horses (group B) than those found in the apparently healthy horses (group A). The observed neutropaenia could be a consequent of overwhelming bacterial infection or severe chronic inflammation which characterizes equine laminitis. This result is comparable to but slightly different from the findings of Riber et al., (1995) who documented lower number of neutrophils in acute laminitis horses compared to the control. Severe inflammatory diseases, regardless of inciting agent, can lead to neutropenia if neutrophil margination into inflamed tissues exceeds the release of neutrophils from the bone marrow (Steven, 2000). With small storage pool of segmented neutrophils in the equine marrow, it is common for horses with overwhelming bacterial infection to have neutropenia (Steven, 2000). Lymphocytes, monocytes, basiophils and eosinophils remained statistically invariable (Table 2). The mean serum biochemical indices such as AST, ALT, ALP, total protein (TP), albumin, globulin and creatinine did not vary significantly (P > 0.05) between groups. Non-significant variation in the enzymatic activities of AST, ALT and ALP could suggest that there may not have the muscular, renal, intestinal and hepatic disorders associated with chronic laminitis in this study. However, Riber et al., (1995), Rashid, (1997), Hussain and Yousaf, (2004) reported a significant increased enzymatic activity in the horses with acute laminitis. Total protein (TP), albumin, globulin and creatinine in horses with laminitis did not differ significantly (p < 0.05) with those found in the apparently healthy Nigerian horses (control). These results were comparable with the report of Ihedioha and Agina (2013) in apparently health Nigerian horses and similar to the findings of Riber et al., (1995) who also reported non-significant (p > 0.05) variation in the mean values of total plasma protein (TPP) between acute laminitis and non-affected Andalusian horses.
In conclusion, this study has documented leucopoenia and neutropaenia in chronic laminitis affected horses and a non-significant enzymatic changes of AST, ALT and ALP were observed in between control and horses with laminitis.

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